• Title/Summary/Keyword: Cell assay

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Effects of Garcinia cambogia Extract on the Adipogenic Differentiation and Lipotoxicity (가르시니아 캄보지아 추출물의 지방세포 분화 및 지방 독성에 미치는 영향)

  • Kang, Eun Sil;Ham, Sun Ah;Hwang, Jung Seok;Lee, Chang-Kwon;Seo, Han Geuk
    • Food Science of Animal Resources
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    • v.33 no.3
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    • pp.411-416
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    • 2013
  • This study aimed to examine the mechanisms underlying the effects of Garcinia cambogia extract on the adipogenic differentiation of 3T3-L1 cells and long-chain saturated fatty acid-induced lipotoxicity of HepG2 cells. 3T3-L1 preadipocytes, mouse embryonic fibroblast-adipose like cell line, were treated with MDI solution (0.5 mM IBMX, 1 ${\mu}M$ dexamethasone, 10 ${\mu}g/mL$ insulin) to generate a cellular model of adipocyte differentiation. Using this cellular model, the anti-obesity effect of Garcinia cambogia extract was evaluated. MDI-induced lipid accumulation and expression of adipogenesis-related genes were detected by Oil red O staining, Nile Red staining, and Western blot analysis. Effects Garcinia cambogia extract on palmitate-induced lipotoxicity was also analyzed by MTT assay, LDH release, and DAPI staining in HepG2 cells. Garcinia cambogia extract significantly suppressed the adipogenic differentiation of preadipocytes and intracellular lipid accumulation in the differentiating adipocytes. Garcinia cambogia extract also markedly inhibited the expression of peroxisome proliferator- activated receptor ${\gamma}2$ ($PPAR{\gamma}2$), CCAT/enhancer-binding protein ${\alpha}$ ($C/EBP{\alpha}$), and adipocyte protein aP2 (aP2). In addition, Garcinia cambogia extract significantly attenuated palmitate-induced lipotoxicity in HepG2 cells. Palmitateinduced cellular damage and reactive aldehydes were also significantly reduced in the presence of Garcinia cambogia extract. These findings suggest that the Garcinia cambogia extract inhibits the adipogenic differentiation of 3T3-L1 preadipocytes, probably by regulating the expression of multiple genes associated with adipogenesis such as $PPAR{\gamma}2$, $C/EBP{\alpha}$, aP2, and thereby modulating fatty acid-induced lipotoxicity to reduce cellular injury in hepatocytes.

Clarithromycin Therapy for Scrub Typus (쯔쯔가무시(Tsutusgamushi)병에서 Clarithromycin의 치료 효과)

  • Kim, Soon;Jung, Eun Mi;Moon, Kyung Hyun;Yoe, Sung Yeob;Eum, Soo Jung;Lee, Joo Hyung;Jo, Sung Rae;Ma, Sang Hyuk
    • Pediatric Infection and Vaccine
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    • v.9 no.2
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    • pp.175-181
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    • 2002
  • Purpose : Scrub typhus(tsutsugamushi disease) is a febrile disease characterized by fever, rash, eschar, lymphadenopathy. Therapy with tetracycline(doxycycline) or chloramphenicol is currently recommended for the treatment for scrub typhus. But there are limitations in usage a tetracycline(doxycycline) for scrub typhus in the children. Recently, there was a report that azithromycin, a macrolide antibiotic was used for scrub typhus in pregnant woman successfully. So we evaluated the effectiveness of the Clarithromycin, other a macrolide antibiotic, for scrub typhus. Methods : Seven patients with scrub typhus at department of internal medicine and three patients with scrub typhus at department of pediatrics Masan Fatima Hospital were involved for this study. A serologic diagnosis for scrub typhus were performed by use of passive hemagglutination test. Clarithromycin(Abbott Laboratories, North Chicago, IL, USA) was administrated orally in a daily dose of 500 mg for adult patients and 15 mg/kg/bid/day for pediatric patients. Results : There were 7 cases of adult patients, varying from 28 to 76 years of age and 3 cases of pediatirc patients, varying from 4 to 7 years of age with scrub typhus. All of cases had fever, myalgia, headache, rash, eschar. Seven cases had positive passive hemagglutination test and eight cases had abnormal liver function. Mean duration for the removal of fever after medication was 1.3 day(1~2 days) and all cases were recovered without complications. Conclusion : Our results suggest that Clarithromycin therapy may be effective for scrub typhus.

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Diagnostic Value of Serum Procalcitonin in Febrile Infants Under 6 Months of Age for the Detection of Bacterial Infections (발열이 있는 6개월 미만의 영아에서 세균성 감염에 대한 procalcitonin의 진단적 가치)

  • Kim, Nam Hyo;Kim, Ji Hee;Lee, Taek Jin
    • Pediatric Infection and Vaccine
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    • v.16 no.2
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    • pp.142-149
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    • 2009
  • Purpose : The aim of this study was to determine the diagnostic value of serum procalcitonin (PCT) compared with that of C-reactive protein (CRP) and the total white blood cell count (WBC) in predicting bacterial infections in febrile infants<6 months of age. Methods : A prospective study was performed with infants <6 months of age who were admitted to the Department of Pediatrics with a fever of uncertain source between July and September 2008. Spinal taps were performed according to clinical symptoms and physical examination. Serum PCT levels were measured using an enzyme-linked fluorescent assay. Results : Seventy-one infants (mean age, 2.62 months) were studied. Twenty-six infants (36.6%) had urinary tract infections (UTIs), and 22 infants (31.0%) had viral meningitis. The remaining infants had acute pharyngitis (n=1), herpangina (n=1), upper respiratory tract infections (n=7), acute bronchiolitis (n=8), acute gastroenteritis (n=4), and bacteremia (n=2). The median WBC and CRP levels were significantly higher in infants with UTIs than in infants with viral meningitis. However, there were no differences in the median PCT levels between the groups (0.14 ng/mL vs. 0.11 ng/mL, P=0.419). The area under the receiver operating characteristic curve was 0.792 (95% CI, 0.65-0.896) for WBC, 0.77 (95% CI, 0.626-0.879) for CRP, and 0.568 (95% CI, 0.417-0.710) for PCT. An elevated WBC count (>11,920/${\mu}L$) and an increased CRP level (>1.06mg/dL) were significant predictors of UTIs based on multiple logistic regression analysis. Conclusion : Serum PCT concentrations should be interpreted with caution in infants <6 months of age with a fever of uncertain source.

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Introduction and Expression of PAP gene using Agrobacterium in Scrophularia buergeriana Miquel (Agrobacterium을 이용한 PAP 유전자의 현삼으로 도입 및 형질발현)

  • Yu, Chang-Yeon;Seong, Eun-Soo;Lim, Jung-Dae;Huang, Shan-Ai;Chae, Young-Am
    • Korean Journal of Medicinal Crop Science
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    • v.9 no.2
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    • pp.156-165
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    • 2001
  • Exogeneous application of pokeweed antiviral protein (PAP), a ribosomal-inacivating protein in the cell wall of Phytolacca americana (pokeweed) protects heterologous plants from viral and fungal infection. A cDNA clone of PAP introduced into Scrophularia buergeriana Miquel by thransformation with Agrobacterium tumefaciences. For plant transformation, explants were precultured on shoot induction medium without kanamycin for 2-5 day, and then they were cocultured with Agrobacterium for 10 minutes. The explants were placed on co culture medium in dark condition, $28^{\circ}C$ for 2days. After explants were washed in MS liquid medium, they were transferred into selection medium including kanamycin 50mg/L (MS salts+1mg/ l BAP+2mg/ l TDZ+0,2mg/ l NAA+MS vitamin+3% sucrose+0.8% agar, pH5.8). From PCR analysis, NPT II band was confirmed in transgenic plant genome and showed resistance against fungi in antifungal activity test. Micro assay to which protein extracted from transgenic line were added, revealed hyphae growth inhibition and no spore germination at high concentration. The characteristics of inhibited hyphae was represented transparent and thin. Expression of PAP in transgenic plants offers the possibility of developing resistance to viral and fungal infection.

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Variations in antioxidant activity in Protaetia brevitarsis larvae depending on the feeding source (먹이원에 따른 흰점박이꽃무지(Protaetia brevitarsis) 유충의 항산화활성)

  • Kim, Hye Soo;Park, Hyun-Young;Kwon, Hyun-Sook;Lee, Sang-Ho;Ha, Jun;Lee, Sang-Won;Cho, Soo-Jeong
    • Journal of Mushroom
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    • v.17 no.4
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    • pp.261-267
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    • 2019
  • The objective of this study was to evaluate the antioxidant activity of extracts of Protaetia brevitarsis larvae fed on fermented oak sawdust (FOS) or spent mushroom substrates (SMS, Pleurotus eryngii). Total polyphenol content was 32% higher in extracts of larvae fed on SMS (P. eryngii) (75.33±0.43 mg GAE/g) than in extracts of larvae fed on FOS (57.02±1.73 mg GAE/g). The flavonoid content of extracts of larvae grown on FOS and SMS (P. eryngii) was 24.6±0.28 mg/g and 25.4±0.75 mg/g, respectively. DPPH radical scavenging activity increased in an extract concentration-dependent manner, and the DPPH radical scavenging capacity of the extract of larvae produced on SMS (P. eryngii) was higher than that of the larvae produced on FOS. The reducing power of the larval extracts produced on FOS and SMS (P. eryngii) increased in an extract concentration-dependent manner, but there was no significant difference between them. The extract of larvae fed on SMS (P. eryngii) (66.55±0.99 uM TE/g) had a higher oxygen radical absorbance capacity (ORAC) than extracts of larvae grown on FOS (76.32±0.48 uM TE/g). The effect of larval extracts on cell proliferation was investigated using a WST-1 (4-[3-(4-iodophenyl)-2-(4-nitrophenyl)-2H-5-tetrazolio]-1,3-benzene disulfonate) assay on RAW 264.7 cells. When cells were treated with larval extracts produced on FOS and SMS (P. eryngii) at concentrations of 0, 2, 4, 8, 16, 32, 40, and 64 mg/ml, RAW 264.7 cells proliferated at 90% or more. Therefore, larval extracts produced on FOS and SMS (P. eryngii) were not toxic to RAW 264.7 cells.

Detection of Campylobacter jejuni in food and poultry visors using immunomagnetic separation and microtitre hybridization

  • Simard, Ronald-E.
    • Proceedings of the Korean Society of Fisheries Technology Conference
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    • 2000.05a
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    • pp.71-73
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    • 2000
  • Campylobacter jejuni is most frequently identified cause of cause of acute diarrhoeal infections in developeed countries, exceeding rates of illness caused by both salmonella and shigilla(Skirrow, 1990 ; Lior 1994). Previous studies on campylobacter jejuni contamination of commercial broiler carcasses in u.s.(Stern, 1992). Most cases of the disease result from indirect transmission of Campylobactor from animals via milk, water and meat. In addition to Campylobactor jejuni. the closely relates species Campylobactor coli and Campylobactor lari have also been implicated as agents of gastroenteritis in humans. Campylobactor coli represented only approximately 3% of the Campylobactor isolates from patients with Campylobactor enteritis(Griffiths and Park, 1990) whereas Campylobactor coli is mainly isolated from pork(Lmmerding et al., 1988). Campylobactor jejuni has also been isolated from cases of bacteremia, appendicitis and, recently, has been associated with Guillai-Barre syndrome(Allos and Blaser, 1994; von Wulffen et al., 1994; Phillips, 1995). Studies in volunteers indicated that the infectious dose for Campylobactor jejuni is low(about 500 organisms)(Robinson, 1981). The methods traditionally used to detect Campylobactor ssp. in food require at least two days of incubation in an enrichment broth followed by plating and two days of incubation on complex culture media containing many antibiotics(Goossens and Butzler, 1992). Finnaly, several biochemical tests must be done to confirm the indentification at the species level. Therfore, sensitive and specific methods for the detection of small numbers of Campylobactor cells in food are needed. Polymerase chain reaction(PCR) assays targeting specific DNA sequences have been developed for the detection of Campylobactor(Giesendorf and Quint, 1995; Hemandex et al., 1995; Winter and Slavidk, 1995). In most cases, a short enrichment step is needed to enhance the sensitivity of the assay prior to detection by PCR as the number of bacteria in the food products is low in comparison with those found in dinical samples, and because the complex composition of food matrices can hinder the PCR and lower its sensitivity. However, these PCR systems are technically demanding to carry out and cumbersome when processing a large number of samples simutaneously. In this paper, an immunomagnetic method to concentrate Campylobactor cells present in food or clinical samples after an enrichment step is described. To detect specifically the thermophilic Campylobactor. a monoclonal antibody was adsorbed on the surface of the magnetic beads which react against a major porin of 45kDa present on the surface of the cells(Huyer et al., 1986). After this partial purification and concentration step, detection of bound cells was achieved using a simple, inexpensive microtitre plate-based hybridization system. We examined two alternative detection systems, one specific for thermophilic Campylobactor based on the detection of 23S rRNA using an immobilized DNA probe. The second system is less specific but more sensitive because of the high copy number of the rRNA present in bacterial cell($10^3-10^4$). By using specific immunomagnetic beads against thermophilic Campylobactor, it was possible to concentrate these cells from a heterogeneous media and obtain highly specific hybridization reactions with good sensitivity. There are several advantages in using microtitre plates instead of filter membranes or other matrices for hybridization techniques. Microtitre plates are much easier to handle than filter membranes during the adsorption, washing, hybridization and detection steps, and their use faciilitates the simultanuous analysis of multiple sample. Here we report on the use of a very simple detection procedure based on a monoclonal anti-RNA-DNA hybrid antibody(Fliss et al., 1999) for detection of the RNA-DNA hybrids formed in the wells.

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Short-term changes of phytoplankton communities after nutrient addition and establishment of stable mass culture condition to prepare the type approval test of USCG Phase-II in mesocosm enclosure (메소코즘에서 USCG phase-II 형식승인 대비 영양염 첨가에 따른 식물플랑크톤 대량 배양조건 확립 및 군집구조의 단주기변화)

  • Baek, Seung Ho;Lee, Min Ji;Shin, Kyoungsoon
    • Journal of the Korea Academia-Industrial cooperation Society
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    • v.17 no.4
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    • pp.34-42
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    • 2016
  • In order to prepare for the type approval test for the United States Coast Guard (USCG) Phase-II of Ballast Water Treatment System (BWTS), a phytoplankton mass culture was conducted in a mesocosm enclosure. We evaluated the response of the phytoplankton community after nutrient addition (+N, +P, and +NP) and investigated the development of the species with increasing culture time. After nutrient dosing, the phytoplankton population significantly (p < 0.05) increased from day 1 to day 3, depending on the nutrient treatments In particular, the specific growth rate of the phytoplankton community in the case of +NP treatment and + N treatment were estimated to be $2.47d^{-1}$ and $1.98d^{-1}$, respectively. The phytoplankton population density in the case of + NP treatment was approximately 50 times higher than that of the control group, suggesting that these treatments could be useful for mass culturing phytoplankton (> 75% of natural community) for the approval regulation of USCG Phase-II. In the phytoplankton community of the mesocosm, Pseudo-nitzchia spp. dominated in the logarithmic growth phase. The cell density decreased significantly (p < 0.05) with increasing time, coinciding with the nutrient limitation. At that time, the dominance of Pseudo-nitzchia spp. shifted to that of Cylindrotheca closterium. Therefore, the optimum nutrient concentration ($N:30{\mu}M$, $P:3{\mu}M$) and reasonable harvesting time (after 3 days in summer) found in this study for the mass culturing of phytoplankton may be helpful to meet the USCG Phase-II biological criteria to be used in BWTS.

Characterization and Cloning of the Gene Encoding Autoregulator Receptor Protein from Streptomyces longwoodensis (Streptomyces longwoodensis로부터 Autoregulator Receptor Protein 유전자의 클로닝 및 특성)

  • Yeo Soo-Hwan;Lee Sung-Bong;Kim Hyun-Soo
    • Microbiology and Biotechnology Letters
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    • v.33 no.2
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    • pp.96-105
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    • 2005
  • For screening of autoregulator receptor gene from Streptomyces longwoodensis, PCR was performed with primers of receptor gene designed on the basis of amino acid sequences of autoregulator receptor proteins with known function. PCR products were subcloned into the BamHIsite of pUC19 and transformed into the E. coli $DH5{\alpha}$. The isolated plasmid from transformant contained the fragment of 100 bp, which was detected on $2\%$ gel after BamHI treatment. The insert, 100 bp PCR product, was confirmed as the expected internal segment of gene encoding autoregulator receptor protein by sequencing. Southern and colony hybridizations with the 100 bp fragment as a probe allowed to select a genomic clone of S. longwoodensis, pSLT harboring a 4.4 kb SphI fragment. Nucleotide sequencing analyses revealed a 651 bp open reading frame(ORF) were isolated protein showing moderate homology ($35{\sim}46\%$) with the ${\Gamma}$-butyrolactone autoregulator receptors from Streptomyces sp., and this ORF was named sltR The sltR/pET-17b plasmid was constructed to overexpress the recombinant SltR protein (rSltR) in E. coli BL21 (DE3)/pLysS, and the rSltR protein was purified to homogeneity by DEAE-Sephacel column chromatography, and DEAE-5PW chromatography (HPLC). The molecular mass of the purified rSltR protein was 55 kDa by HPLC gel-filtration chromatography and 28 kDa by SDS-PAGE, indicating that the rSltR protein is present as a dimer. A binding assay with tritium-labeled autoregulators revealed that the rSltR has clear binding activity with a A-factor type autoregulator as the most effective ligand.

Apoptosis and Development of Porcine Parthenogenetic Embryos Activated and Cultured in Different Condition (활성화 및 배양조건이 돼지 단위발생란의 발달 및 Apoptosis에 미치는 영향)

  • Hwang In-Sun;Seo Jin-Sung;Cheong Hee-Tae;Im Gi-Sun
    • Reproductive and Developmental Biology
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    • v.30 no.1
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    • pp.65-70
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    • 2006
  • This study investigated apoptosis and in vitro development of parthenogenetic preimplantation porcine embryos. In vitro matured oocytes for $42{\sim}44h$ were used. Apoptotic cell death was analyzed by using a terminal deoxynucleatidyl transferase mediated deoxyuridine 5-triphosphate nick-end tabling (TUNEL) assay. In experiment 1, oocytes were activated with two electric pulses (CH) of 1.2 kV/cm for $30{\mu}sec$ (E), E + 6-dimethylaminopurine (6-DMAP) or E + cycloheximide (CH) and cultured in PZM-3 under 5% $CO_2$ in air at $38.5^{\circ}C$. In experiment 2, oocytes were activated by E and cultured in PZM-3 or NCSU-23 under a gas atmosphere of 20% $O_2$ ($5%\;CO_2$, in air) or 5% $O_2$ $(5%\;CO_2,\;5%\;O_2\;90%\;N_2)\;at\;38.5^{\circ}C$. Oocytes activated with E+6-DMAP or E+CH showed higher blastocyst rates (36.3% and 32.5%) compared to E alone (27.7%). The frequency of apoptosis according to treatments were 5.3%, 7.7% and 7.1% respectively. Oocytes activated with E alone showed lower (P<0.05) frequency of apoptosis compared to other groups. In experiment 2, parthenotes cultured in PZM-3 showed slightly higher blastocyte rates (28.2% and 29.7%) compared to NCSU-23 (22.6% and 24.4%) regardless of atmosphere. Blastocysts generated in PZM-3 showed lower (P<0.05) apoptosis rate under 20% $O_2$ (9.2% vs 16.9%), whereas those in NCSU-23 had slightly lower apoptosis rate under 5% $O_2$ (14.0% vs 18.4%). This result represents that activation method and culture condition could affect the frequency of apoptosis as well as in vitro developmental rate.

Treatment of Reproductive Dysfunctions and Reproductive Monitoring Using Ultrasonography in Dairy Cow (초음파 검사에 의한 젖소 번식 검진과 번식 장애 치료)

  • Lim, W.H.;Oh, K.S.;Seo, G.J.;Hwang, S.S.;Kim, B.S.;Bae, C.S.;Kim, S.H.;Kim, J.T.;Park, I.C.;Park, S.G.;Son, C.H.
    • Journal of Embryo Transfer
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    • v.21 no.3
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    • pp.217-223
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    • 2006
  • This study was carried out principally to obtain the basic data for the improvement of the reproductive performance and production using plasma progesterone assay and ultrasonography in dairy cow. The results obtained from this studies were as follows. The results of reproductive examination in 85,983 cows were ovarian diseases 40,399 (47.0%), uterine diseases 11,912 (13.9%), pregnancy or pregnant failures 26,587 (30.9%), adhesion of reproductive tracts 172 (0.2%), freemartin 8 (0.01%), and others 6,905 (8.3%), respectively. The treatment status of reproductive dysfunction in 30,241 cows were silent heat or error of estrus detection 14,909 (49.3%), follicular cysts 3,750 (12.4%), luteal cysts 907 (3.0%), inactive ovaries 665 (2.2%), granulosa cell tumor of ovary 3 (0.01%) and endometritis 6,986 (23.1%), respectively. The indices of reproductive efficiency after the periodical examination of reproductive status were as follows; the mean intercalving inteual was reduced from 475 days at the first examination to 381 days at the last examination of reproductive status, the mean interval calving to conception was reduced from 186 to 98 days, the mean interval calving to first service was reduced from 106 to 66 days, the cows showing heat by 60 days postpartum were increased from 32 to 90%, the mean conception rate to first service was increased from 42 to 64%, and the mean service per conception was reduced from 2.6 to 1.8 times, respectively.